Acquired melanocyte stimulating hormone-inducible chemotaxis following macrophage fusion with Cloudman S91 melanoma cells.
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Fusion of Cloudman S91 melanoma cells with macrophages results in hybrids with increased metastatic potential. Here, we report that such hybrids acquire new pathways for motility. Compared to parental melanoma cells and low metastatic hybrids, the metastatic hybrids showed far stronger responses to 3T3- and lung fibroblast-conditioned media, primary lung slices, fibronectin (FN), and a Mr 120,000 FN fragment and, unlike parental cells, were further stimulated by pretreatment with melanocyte-stimulating hormone/1-methyl-3-isobutylxanthine. Hybrid migration was due primarily to chemotaxis, with chemokinesis being a minor component. Thus, the metastatic hybrids acquired melanocyte-stimulating hormone-inducible motility, perhaps reflecting the FN fragment chemotaxis of macrophages. The results support a long-standing hypothesis that metastasis is initiated following hybridization between tumor-invading phagocytes and cells of the primary tumor. (+info)
Mouse semaphorin H induces PC12 cell neurite outgrowth activating Ras-mitogen-activated protein kinase signaling pathway via Ca(2+) influx.
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We recently showed that mouse semaphorin H (MSH), a secreted semaphorin molecule, acts as a chemorepulsive factor on sensory neurites. In this study, we found for the first time that MSH induces neurite outgrowth in PC12 cells in a dose-dependent manner. Comparison of Ras-mitogen-activated protein kinase (MAPK) signaling pathways between MSH and nerve growth factor (NGF) revealed that these pathways are crucial for MSH action as well as NGF. K-252a, an inhibitor of tyrosine autophosphorylation of tyrosine kinase receptors (Trks), did not inhibit the action of MSH, suggesting that MSH action occurs via a different receptor than NGF. L- and N-types of voltage-dependent Ca(2+) channel blockers, diltiazem and omega-conotoxin, inhibited MSH-induced neurite outgrowth and MAPK phosphorylation in a Ca(2+)-dependent manner. A transient elevation in intracellular Ca(2+) level was observed upon MSH stimulation. These findings suggest that extracellular Ca(2+) influx, followed by activation of the Ras-MAPK signaling pathway, is required for MSH induced PC12 cell neurite outgrowth. (+info)
Evidence that melanocortin 4 receptor mediates hemorrhagic shock reversal caused by melanocortin peptides.
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Melanocortin peptides are known to be extremely potent in causing the sustained reversal of different shock conditions, both in experimental animals and humans; the mechanism of action includes an essential brain loop. Three melanocortin receptor subtypes are expressed in brain tissue: MC(3), MC(4,) and MC(5) receptors. In a volume-controlled model of hemorrhagic shock in anesthetized rats, invariably causing the death of control animals within 30 min after saline injection, the i.v. bolus administration of the adrenocorticotropin fragment 1-24 (agonist at MC(4) and MC(5) receptors) at a dose of 160 microg/kg i.v. (54 nmol/kg) produced an almost complete and sustained restoration of cardiovascular and respiratory functions. An equimolar dose of gamma(1)-melanocyte stimulating hormone (selective agonist at MC(3) receptors) was completely ineffective. The selective antagonist at MC(4) receptors, HS014, although having no influence on cardiovascular and respiratory functions per se, dose-dependently prevented the antishock activity of adrenocorticotropin fragment 1-24, with the effect being complete either at the i.v. dose of 200 microg/kg or at the i.c.v. dose of 5 microg/rat (17-20 microg/kg). We concluded that the effect of melanocortin peptides in hemorrhagic shock is mediated by the MC(4) receptors in the brain. (+info)
Model studies of enzymatic NH2-terminal acetylation of porteins with des-Nalpha1-acetyl-alpha-melanotropin as a substrate.
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The present study describes the acetylation by an enzyme present in calf lens of a synthetic tridecapeptide [analogous to alpha-melanotropin (alpha-melanocyte stimulating hormone) but lacking the naturally occurring NH2-terminal acetyl group: des-Nalpha1-Ac-alpha-melanotropin]. The reaction is specific for the alpha-amino group of the NH2-terminal amino acid. The minimum length required for the substrate to become acetylated appears to be a sequence of five to eight amino acid residues. Modification of the internal lysine decreases the incorporation of acetate, irrespective of the size of the blocking group. (+info)
Contribution of changes in the chloride driving force to the fading of I(GABA) in frog melanotrophs.
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Chloride redistribution during type A gamma-aminobutyric acid (GABA(A)) currents (I(GABA)) has been investigated in cultured frog pituitary melanotrophs with imposed intracellular chloride concentration ([Cl(-)](i)) in the whole cell configuration or with unaltered [Cl(-)](i) using the gramicidin-perforated patch approach. Prolonged GABA exposures elicited reproducible decaying currents. The decay of I(GABA) was associated with both a transient fall of conductance (g(GABA)) and shift of current reversal potential (E(GABA)). The shift of E(GABA) appeared to be time and driving force dependent. In the gramicidin-perforated patch configuration, repeated GABA exposures induced currents that gradually vanished. The fading of I(GABA) was due to persistent shifts of E(GABA) as a result of g(GABA) recovering from one GABA application to another. In cells alternatively clamped at potentials closely flanking resting potential and submitted to a train of brief GABA pulses, a reversal of I(GABA) was observed after 150 s recording. It is demonstrated that, in intact frog melanotrophs, shifts of E(GABA) combine with genuine receptor desensitization to depress I(GABA). These findings strongly suggest that shifts of E(GABA) may act as a negative feedback, reducing the bioelectrical and secretory responses induced by an intense release of GABA in vivo. (+info)
Widespread expression of Agouti-related protein (AGRP) in the chicken: a possible involvement of AGRP in regulating peripheral melanocortin systems in the chicken.
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Agouti-related protein (AGRP) is a naturally occurring antagonist of melanocortin action. It is expressed mainly in the arcuate nucleus where it plays an important role in the hypothalamic control of feeding and energy homeostasis by antagonism of central melanocortin 4 receptors in mammals. Besides in the brain, the melanocortin 4 receptor is expressed in numerous peripheral tissues in the chicken. To examine whether or not the peripheral melanocortin 4 receptor signaling could be regulated by AGRP, we cloned and localized the expression of the AGRP gene in the chicken. The chicken AGRP gene was found to encode a 154 or 165 amino acid protein, depending on the usage of two alternative translation initiation sites. The coding sequence consisted of three exons, like that of mammalian species. The C-terminal cysteine-rich region of the predicted AGRP displayed high levels of identity to mammalian counterparts (78-84%) and all 10 cysteine residues conferring functional conformation of AGRP were conserved; however, other regions showed apparently no homology, suggesting that biological activities of AGRP are located in its C-terminal region. RT-PCR analysis detected the AGRP mRNA in all tissues examined: the brain, adrenal gland, heart, liver, spleen, gonads, kidney, uropygial gland, skeletal muscle and adipose tissues. Interestingly, the skin also expressed the AGRP mRNA, where Agouti, another melanocortin receptor antagonist regulating hair pigmentation, is expressed in rodents. Most of those AGRP-expressing tissues have been demonstrated to express melanocortin 4 receptors and/or other subtypes of melanocortin receptor whose mammalian counterparts can bind AGRP. These results imply the possibility that some peripheral melanocortin systems could be regulated by the functional interaction between melanocortins and AGRP at melanocortin receptors in the chicken. (+info)
Pathophysiological role of leptin in obesity-related hypertension.
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To explore the pathophysiological role of leptin in obesity-related hypertension, we examined cardiovascular phenotypes of transgenic skinny mice whose elevated plasma leptin concentrations are comparable to those seen in obese subjects. We also studied genetically obese KKA(y) mice with hyperleptinemia, in which hypothalamic melanocortin system is antagonized by ectopic expression of the agouti protein. Systolic blood pressure (BP) and urinary catecholamine excretion are elevated in transgenic skinny mice relative to nontransgenic littermates. The BP elevation in transgenic skinny mice is abolished by alpha(1)-adrenergic, beta-adrenergic, or ganglionic blockers at doses that do not affect BP in nontransgenic littermates. Central administration of an alpha-melanocyte-stimulating hormone antagonist causes a marked increase in cumulative food intake but no significant changes in BP. The obese KKA(y) mice develop BP elevation with increased urinary catecholamine excretion relative to control KK mice. After a 2-week caloric restriction, BP elevation is reversed in nontransgenic littermates with the A(y) allele, in parallel with a reduction in plasma leptin concentrations, but is sustained in transgenic mice overexpressing leptin with the A(y) allele, which remain hyperleptinemic. This study demonstrates BP elevation in transgenic skinny mice and obese KKA(y) mice that are both hyperleptinemic, thereby suggesting the pathophysiological role of leptin in some forms of obesity-related hypertension. (+info)
Mahogany (1377-1428) enters brain by a saturable transport system.
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The mouse mahogany gene encodes a protein that is involved in the suppression of diet-induced obesity. We studied the ability of its widely conserved C-terminal fragment to cross the blood-brain barrier (BBB) in mice. Multiple-time regression analysis showed that the entry rate (K(i)) of (125)I-mahogany (1377-1428) from blood-to-brain was 5.5 x 10(-4) ml/g. min. After coinjection of unlabeled mahogany (1377-1428), the K(i) was significantly decreased, showing the self-inhibition characteristic of a saturable transport mechanism. The excess mahogany (1377-1428) did not change the influx rate of (99m)Tcalbumin, the vascular control, indicating a lack of disruption of the BBB. Statistically significant cross-inhibition was not seen with agouti-related protein (83-132), melanin-concentrating hormone, epidermal growth factor, leptin, a melanocortin-4 receptor antagonist, or alpha-melanocyte-stimulating hormone. HPLC showed that most of the injected (125)I-mahogany (1377-1428) reached the brain intact, and capillary depletion with washout showed that most of it reached the parenchyma. There was no brain-to-blood efflux system for mahogany (1377-1428) but rather retention after i.c.v. administration, and the octanol/buffer partition coefficient showed low lipophilicity. Thus, the results show that the C-terminal peptide product encoded by the mahogany gene crosses the BBB by a transport mechanism that is saturable. The ability of this system to be regulated indicates the therapeutic potential of mahogany (1377-1428) in the treatment of obesity. (+info)